Crystal Structure Analysis of Phosphatidylcholine-GM2-Activator Product Complexes: Evidence for Hydrolase Activity

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• 作者：Christine S. Wright ; Li-Zhi Mi ; Sangderk Lee ; and Fraydoon Rastinejad
• 刊名：Biochemistry
• 出版年：2005
• 出版时间：October 18, 2005
• 年：2005
• 卷：44
• 期：41
• 页码：13510 - 13521
• 全文大小：6747K
• 年卷期：v.44,no.41(October 18, 2005)
• ISSN：1520-4995

文摘

GM2-activator protein (GM2AP) is a lysosomal lipid transfer protein with important biologicalroles in ganglioside catabolism, phospholipid metabolism, and T-cell activation. Previous studies of crystalstructures of GM2AP complexed with the physiological ligand GM2 and platelet activating factor (PAF)have shown binding at two specific locations within the spacious apolar pocket and an ordering effect ofendogenous resident lipids. To investigate the structural basis of phospholipid binding further, GM2APwas cocrystallized with phosphatidylcholine (PC), known to interact with GM2AP. Analysis of threecrystal forms revealed binding of single chain lipids and fatty acids only and surprisingly not intact PC.The regions of best defined electron density are consistent with the presence of lyso-PC and oleic acid,which constitute deacylation products of PC. Their acyl tails are in stacking contact with shorter, lesswell-defined stretches of electron density that may represent resident fatty acids. The GM2AP associatedhydrolytic activity that generates lyso-PC was further confirmed by mass spectrometry and enzymaticassays. In addition, we report the structures of (i) mutant Y137S, assessing the role of Tyr¹³⁷ in lipidtransfer via the hydrophobic cleft, and (ii) apo-mouse GM2AP, revealing a hydrophobic pocket with aconstricted opening. Our structural results provide new insights into the biological functions of GM2AP.The combined effect of hydrolytic and lipid transfer properties has profound implications in cellularsignaling.